**5. NPs for delivering DNA and mRNA vaccines**

Using DNA and mRNA for intracellular production of oncogenic proteins or peptides as tumor Ags becomes an attractive strategy for developing cancer vaccines thanks to the advances in biotechnology which allows gene encoding proteins of interest to be easily manufactured in batch and be further modified with nucleic acid sequences that encode for proteins with immunostimulatory functions, for example, flagellin and a toll-like receptor 5 agonist (TLR5a). Unfortunately, previous clinical trials on DNA cancer vaccines, majority of which were administered as naked DNA via the intramuscular route, showed generally poor response rates, despite employment of viral vectors and electroporation able to improve the transfection of DNA vaccines, both of which cause safety and compliance concerns [61, 62]. Alternatively, NPs engineered as a VADS for intracellular delivery of DNA and mRNA provide a promising strategy for developing nucleotide-based cancer vaccines and possess several advantages [6]: (1) synthetic material-constituted NPs are safer than viral vectors, (2) NPs can stabilize and protect gene therapeutics from nuclease-mediated degradation [63], (3) DNA- and RNA-loaded NPs can be administered by injection-free tools, such as microneedles for non-parenteral delivery [64], and (4) nanocarriers can be easily modified with targeting moieties, for example, mannose, to achieve DC-targeted delivery and transfection [14, 65].

It is reported that cationic liposomes and lipid nanoparticles containing mRNA coding for the tumor-associated Ags gp100 and TRP2 could induce a strong CD8+ T cell activation after a single immunization and treatment of B16F10 melanoma tumors with the mRNAcarried cationic liposomes resulted in tumor shrinkage and extended the overall survival of the treated mice, all of which could be further increased by the combinatory incorporation of the adjuvant LPS, showing the cationic liposomes a promising vector for mRNA vaccine delivery that is capable of inducing a strong cytotoxic T cell response for cancer immunotherapy [66]. Nevertheless, nucleotide-based vaccines, including DNA and mRNA vaccines with their intracellular Ag synthesis, have been shown to be potent activators of a cytotoxic T cell response which is an important prerequisite for successful immunotherapy against many viral diseases and tumors [67], though intracellular delivery of mRNA vaccines to the cytosol of APCs is still not sufficiently well understood and remains somewhat a challenge to clinical translation for cancer immunotherapy [68].
